PROJECT 3: TRANSDUCTION OF HEMATOPOIETIC STEM CELLS FOR ENHANCED IMMUNOTHERAPY OF MELANOMA. Project Leaders: Donald B. Kohn, James S. Economou, Antoni Ribas, Jerome A. Zack ABSTRACT Immunotherapy can be performed by expressing in T cells the genes encoding a T cell receptor (TCR) reactive against a tumor-associated antigen. This Project takes this approach, which is explored in Project 1, further by examining whether administration of hematopoietic stem cells (HSC) transduced with the genes encoding an anti-tumor TCR can lead to de novo production of T cells expressing the introduced TCR genes.. Theoretically, transduced HSC could serve as an additional long-term source of engineered T cells for sustained anti-tumor activity, to augment that of transduced mature T cells. We will develop and perform a Phase I clinical trial to assess the safety and feasibility of administering autologous CD34+ HSC transduced to express a TCR recognizing a peptide from MART-1 for immunotherapy of patients with advanced melanoma. These engineered CD34+ cells will be co-administered along with mature T cells transduced to express the same MART-1 TCR, but with a distinguishable reporter gene. This dual marking approach will allow identification, isolation and characterization of the TCR-expressing T cells produced in vivo from the CD34+ cells and determination of their in vivo biodistribution by using PET imaging. During the planning stage of the clinical trial, we will use pre-clinical models of human T cell production from HSC to perform detailed characterization of the T cells derived from transduced human HSC. Ongoing scientific collaborations will occur with all of the Projects and Cores of this Program. The clinical trial of this Project 3 will be performed in years 4-5 of this program, building on the clinical trial of Project 1 that defines the optimal methods for administration of the TCR-transduced T cells. These studies will assess the potential role of HSC for cancer immunotherapy using TCR-based approaches.